CN105385607A - Lentinus edodes liquid submerged fermentation culture medium formula and fermentation technology - Google Patents
Lentinus edodes liquid submerged fermentation culture medium formula and fermentation technology Download PDFInfo
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- CN105385607A CN105385607A CN201510960502.3A CN201510960502A CN105385607A CN 105385607 A CN105385607 A CN 105385607A CN 201510960502 A CN201510960502 A CN 201510960502A CN 105385607 A CN105385607 A CN 105385607A
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/14—Fungi; Culture media therefor
Abstract
The invention provides a lentinus edodes liquid submerged fermentation culture medium formula. A lentinus edodes liquid submerged fermentation culture medium is prepared from molasses, corn flour, bean pulp flour, magnesium sulfate, monopotassium phosphate, yeast extract and defoaming oil, and the pH value is adjusted to 5.0 before the culture medium is sterilized. The specific technology includes the steps that lentinus edodes slant strains are inoculated into shake flasks containing a liquid culture medium to obtain lentinus edodes liquid strains through culturing; the lentinus edodes liquid strains are transplanted to the liquid culture medium to be cultured; after lentinus edodes mycelium pellets grow over the liquid culture medium, the lentinus edodes liquid strains serving as first-level strains are inoculated into a seed tank with the volume of 500 L and are fermented, then the lentinus edodes strains, serving as second-level strains, in the seed tank with the volume of 500 L are inoculated into a fermentation tank with the volume of 10 m<3> and are fermented, and the lentinus edodes liquid submerged fermentation technology can be completed.
Description
Technical field
The present invention relates to a kind of configuration of mushroom liquid submerged fermentation culture medium and the making method of zymotechnique, belong to biological fermentation field.
Background technology
Mushroom (Lentinulaedodes) is a kind of gill fungus bacterium eating medicine dual-purpose, is one of China's edible medicinal fungus of extensively cultivating.Mushroom contains rich in protein, carbohydrate, fat, electrolytes and minerals, and the medicinal ingredients such as lentinan, terpenoid, steroid simultaneously in mushroom, has the effect such as activating immune system, anti-curing oncoma.
Current mushroom is extensively cultivated in provinces such as China Hubei, Zhejiang, Henan, and big area cultivating champignon needs a large amount of excellent species as support, and current China Lentnus edodes bacterial classification used is still solid spawn, and the production of solid spawn need drop into a large amount of human and material resources.Along with the cross development of each subject, fermentation engineering has been applied to edible mushrooms industry liquid submerged fermentation, compared to solid spawn, liquid submerged fermentation gained liquid spawn is used for edible fungus culturing, having that technological operation is easy, pollution rate is low, send out that bacterium is fast, cell age is consistent, fruiting is neat, low cost and other advantages, is the brand-new route developing Lentnus edodes.Mycelium and the meta-bolites of edible fungus liquid submerged fermentation simultaneously also can be used for extracting effective components.Carry out liquid submerged fermentation it is crucial that whether the formula of liquid nutrient medium used is excellent, whether zymotechnique is applicable to.Current mushroom liquid submerged fermentation culture medium configuration material therefor cost is higher, be not suitable for large-scale commercial production, fermenting process is more common in the shaking flask stage simultaneously, rarely have and carry out in fairly large fermentor tank, when Submerged liquid culturation is to certain scale, the quality of control to whole fermentation of various parameter plays considerable effect, if lab scale (shaking flask or small fermentor) technique is directly overlapped to use in large scale fermentation and often occurs that state modulator is lacked of proper care, and then cause fermentation yield Quality Down.
Summary of the invention:
The application is when preparing fermentation liquid nutrient medium used, to abandon using in laboratory price material costly for the mushroom ferment industrial production of reality, filter out abundance and cheap molasses, Semen Maydis powder and bean cake powder as the carbon source nitrogenous source of mushroom liquid submerged fermentation, add a small amount of magnesium sulfate, mineral element that potassium primary phosphate provides part necessary for mushroom liquid submerged fermentation, add a small amount of yeast extract and provide quick-acting nitrogenous source and somatomedin for fermenting process, add a small amount of anti-crawl agentfoam oil to reduce the generation of foam in fermenting process.The composition of concrete substratum and proportioning are:
Molasses (after process, total sugar concentration is 27%) 5%, Semen Maydis powder 0.3%, bean cake powder 0.3%, yeast extract 0.1%, magnesium sulfate 0.03%, potassium primary phosphate 0.05%, anti-crawl agentfoam oil 0.03%, before medium sterilization, adjust pH is 5.0.
The present invention abandons adopting triangular flask shaking table to cultivate and carries out mushroom liquid culture, only triangular flask shaking table is cultivated the enlarged culturing being used for first class inoculum.After obtaining the first class inoculum of q.s, first class inoculum is transferred in sterile culture inoculation tank, utilizing sterile culture to inoculate tank is pressed in fermentor tank by liquid spawn, avoid and utilize Alcohol Flame ring at fermentation plant, open the inoculation mouth of fermentor tank, the mode that one-level kind in triangular flask pours fermentor tank into is inoculated, greatly can reduce the pollution rate of inoculation like this.Concrete zymotechnique comprises the steps:
(1). by the access of mushroom slant strains containing in the shaking flask of liquid nutrient medium, shaking speed is 160 ~ 180rpm, cultivates 5 ~ 6 days for 24 ~ 26 DEG C; (2). by the mushroom liquid bacterial of gained in step (1) by 2% inoculum size transfer liquid nutrient medium is housed shaking flask in carry out enlarged culturing, shaking speed is 160 ~ 180rpm, cultivates 4 ~ 5 days for 24 ~ 26 DEG C; (3). after the Lentinus Edodes fungus pompon in step (2) covers with liquid nutrient medium, the liquid spawn of gained is transferred in aseptic operating platform sterile culture inoculation tank, coefficient is about 70%; (4). mushroom liquid bacterial sterile culture inoculated in tank is produced in fermentor tank as first order seed access, fermenting process keeps broth temperature 24 ~ 26 DEG C, tank pressure 0.13MPa, stir speed (S.S.) and air flow change with dissolved oxygen, ensure that dissolved oxygen is that 90% ~ 95% (rotating speed controls at 100 ~ 250rpm, and air flow controls at 6 ~ 15m
3/ h), in conjunction with tank body pH Real-Time Monitoring probe display numerical value, adding by stream the pH that saturated sodium carbonate solution controls whole fermenting process fermented liquid is about 4.5.
Ferment 127 hours, and unit volume fermented liquid mycelium pellet volume no longer increases, and pH remains unchanged substantially, and when adding alkali lye without the need to stream, fermentation ends puts tank.In whole fermenting process, sampling in first 24 hours once, after every sampling in 6 hours once, microscopy, survey off-line pH, observe substratum color, hear substratum smell, measure sedimentation mushroom mycelium sphere volume in unit volume fermented liquid after 2 hours simultaneously.
The composition of the composition of the shaking flask described in the present invention and fermention medium and proportioning substratum same as above and proportioning.Unit volume fermented liquid mushroom mycelium sphere volume method of calculation are: pour in 100mL graduated cylinder by the sample mixed after sampling, leave standstill 2 hours, mushroom mycelium sphere volume in Units of Account volume fermented liquid.
The liquid fermenting of mushroom strain, the fermentation level of lab shaker or laboratory 30-50L fermentor tank is also in the world at present in the industry, in producing with regard to international existing needle mushroom, Pleurotus eryngii industrial cultivation, the maximum liquid spawn production technique of liquid fungus seed also only has 200-300L fermentor tank, be not with stirring, the size of uncontrollable ventilation and mycelium pellet, can not carry out fully automatic inoculating.Moreover canister fermentation, time consumption and energy consumption.Labo r usage is large, cost up.
This technique can disposable production 7m
3mushroom liquid bacterial, equipment and process is advanced, and fermentation parameter temperature, potential of hydrogen, ventilation, mixing speed etc. are all controlled, and mycelium pellet size can adjust at any time.Be applicable to batch production automatic production line.
In other formula is produced, mycelial recovery rate is generally 20-30% (fermentating liquid volume ratio) and ferments with this culture medium prescription, and recovery rate is up to 60-70% (fermentating liquid volume ratio).
The culture medium prescription filtered out in the application is suitable for mushroom liquid bacterial submerged fermentation, and the zymotechnique cycle is short, greatly can reduce cost of manufacture and the production cycle of bacterial classification in Lentnus edodes, this beneficial effect can be specialized, magnifies, change in detail.
Embodiment
Embodiment 1
(1). mushroom slant strains being accessed 4 bottles of capacity is that 250mL is equipped with in the shaking flask of 100mL liquid nutrient medium, and shaking speed is 160 ~ 180rpm, cultivates 5 ~ 6 days for 24 ~ 26 DEG C; (2). by the mushroom liquid bacterial of gained in step (1) by 2% inoculum size 4 bottles of capacity of transferring be that 5000mL is equipped with in the shaking flask of 2000mL liquid nutrient medium and carries out enlarged culturing, shaking speed is 160 ~ 180rpm, cultivates 4 ~ 5 days for 24 ~ 26 DEG C; (3). after the Lentinus Edodes fungus pompon in step (2) covers with liquid nutrient medium, the liquid spawn of gained is transferred in aseptic operating platform the sterile culture inoculation tank that volume is 10L, coefficient is about 70%; (4). mushroom liquid bacterial access coefficient sterile culture inoculated in tank is 70% volume is in 500L fermentor tank, inoculum size is about 2%, fermenting process keeps tank temperature 24 ~ 26 DEG C, tank pressure 0.13MPa, stir speed (S.S.) and air flow change with dissolved oxygen, ensure that dissolved oxygen is that 90% ~ 95% (rotating speed controls at 100 ~ 250rpm, and air flow controls at 6 ~ 15m
3/ h), adding by stream the pH that saturated sodium carbonate solution controls whole fermenting process fermented liquid is about 4.5.
Whole fermenting process sampling microscopy hypha form is normal; Along with the carrying out of fermentation, the nutritive substance in substratum is consumed gradually, and the color of substratum becomes bright gradually, and substratum has light mushroom fragrance; Within 65 hours, pH drops to 4.44, starts stream and adds alkali lye by between pH modulation 4.44 to 4.60, and 69 little to add alkali lye up to 112 hours stream frequent, on average per hourly add once, 112 is little of 127 hours, and alkali stream adds frequency and obviously declines, add alkali lye without the need to stream after 127 hours, pH stablizes; Mycelium pellet volume increasing law is: within 36 hours, unit volume fermented liquid mycelium pellet volume starts to increase, within 72 ~ 96 hours, mycelium pellet volume increases fast, 96 ~ 114 hours slowdown in growths, within 126 hours, unit volume fermented liquid mycelium pellet volume reaches maximum value 0.955, has downtrending afterwards.Comprehensive analysis stream adds alkali lye frequency and unit volume fermented liquid mushroom mycelium sphere volume increasing law, 69 ~ 127 hours these one-phases can be summed up as logarithmic phase and the deceleration phase of mushroom liquid fermenting; In conjunction with Lentinus Edodes fungus pompon generation and follow-up cultivating bag inoculation sprouting test result, determine that the best inoculation time of mushroom liquid bacterial is 96 little of 127 hours.
Claims (2)
1. a mushroom liquid submerged fermentation culture medium formula, it is characterized in that, comprise molasses, Semen Maydis powder, bean cake powder, magnesium sulfate, potassium primary phosphate, yeast extract, anti-crawl agentfoam oil, percentage, each composition proportion is: total sugar concentration is the molasses 5% of 27%, Semen Maydis powder 0.3%, bean cake powder 0.3%, yeast extract (nitrogenous source) 0.1%, magnesium sulfate 0.03%, potassium primary phosphate 0.05%, anti-crawl agentfoam oil (biological anti-crawl agentfoam oil PPE) 0.03%, before medium sterilization, adjust pH is 5.0.
2. a mushroom deep liquid fermentation process, is characterized in that, comprises the steps:
(1) by the shaking flask of mushroom slant strains access liquid nutrient medium, shaking speed is 160 ~ 180rpm, cultivates 5 ~ 6 days for 24 ~ 26 DEG C;
(2) by the mushroom liquid bacterial of gained in step (1) by 2% inoculum size transfer liquid nutrient medium is housed shaking flask in carry out enlarged culturing, shaking speed is 160 ~ 180rpm, cultivates 4 ~ 5 days for 24 ~ 26 DEG C;
(3) after the Lentinus Edodes fungus pompon in step (2) covers with liquid nutrient medium, the liquid spawn of gained is transferred in aseptic operating platform sterile culture inoculation tank;
(4) using sterile culture, the mushroom liquid bacterial inoculated in tank accesses fermentor tank as first order seed, fermenting process keeps broth temperature 24 ~ 26 DEG C, tank pressure 0.13MPa, and stir speed (S.S.) and air flow change with dissolved oxygen, rotating speed controls at 100 ~ 250rpm, and air flow controls at 6 ~ 15m
3/ h also ensures that dissolved oxygen is 90% ~ 95%, in conjunction with tank body pH Real-Time Monitoring probe numerical value, adding by stream the pH that saturated sodium carbonate solution controls whole fermenting process fermented liquid is about 4.5, ferment 127 hours, unit volume fermented liquid mycelium pellet volume substantially no longer increases, pH remains unchanged substantially, and when adding alkali lye without the need to stream, fermentation ends puts tank.
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107333982A (en) * | 2017-08-23 | 2017-11-10 | 延边德康生物技术有限公司 | A kind of method for making matrix Bei Lu feeds with soya bean straw using Lenlinus edodes |
| CN107517737A (en) * | 2017-10-20 | 2017-12-29 | 翔天农业开发集团股份有限公司 | A kind of continuous quick bacteria stick manufacture craft of edible mushroom |
| CN107641601A (en) * | 2017-10-20 | 2018-01-30 | 翔天农业开发集团股份有限公司 | A kind of liquid fermentation strain domestication method |
| CN108076973A (en) * | 2018-01-15 | 2018-05-29 | 石家庄学院 | A kind of production method of mushroom concentrated strain |
| CN110583371A (en) * | 2019-09-23 | 2019-12-20 | 贵州同辉食用菌发展有限公司 | Fermentation method of mushroom liquid strain with high activity and high stability |
| CN112703967A (en) * | 2020-12-07 | 2021-04-27 | 贵州大学 | Method for preparing wood rot fungus liquid strain by using yellow serofluid and bean dregs |
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| CN1136393A (en) * | 1996-01-19 | 1996-11-27 | 济南科贝尔生物工程有限公司 | Agricultural chemical for preventing and controlling plant virus prepared by liquid fermentation and its producing technology |
| CN1463578A (en) * | 2002-06-20 | 2003-12-31 | 北京锦绣大地农业股份有限公司 | Submerged culturing method for making mushroom liquid bacterial and culture medium therefor |
| CN104012298A (en) * | 2014-04-29 | 2014-09-03 | 潢川九龙春天农业科技有限公司 | Edible fungus liquid seed submerged fermentation packaged seed production technology and medium formulas thereof |
-
2015
- 2015-12-18 CN CN201510960502.3A patent/CN105385607A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1136393A (en) * | 1996-01-19 | 1996-11-27 | 济南科贝尔生物工程有限公司 | Agricultural chemical for preventing and controlling plant virus prepared by liquid fermentation and its producing technology |
| CN1463578A (en) * | 2002-06-20 | 2003-12-31 | 北京锦绣大地农业股份有限公司 | Submerged culturing method for making mushroom liquid bacterial and culture medium therefor |
| CN104012298A (en) * | 2014-04-29 | 2014-09-03 | 潢川九龙春天农业科技有限公司 | Edible fungus liquid seed submerged fermentation packaged seed production technology and medium formulas thereof |
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107333982A (en) * | 2017-08-23 | 2017-11-10 | 延边德康生物技术有限公司 | A kind of method for making matrix Bei Lu feeds with soya bean straw using Lenlinus edodes |
| CN107517737A (en) * | 2017-10-20 | 2017-12-29 | 翔天农业开发集团股份有限公司 | A kind of continuous quick bacteria stick manufacture craft of edible mushroom |
| CN107641601A (en) * | 2017-10-20 | 2018-01-30 | 翔天农业开发集团股份有限公司 | A kind of liquid fermentation strain domestication method |
| CN107517737B (en) * | 2017-10-20 | 2019-01-08 | 翔天农业开发集团股份有限公司 | A kind of continuous quick bacteria stick manufacture craft of edible mushroom |
| CN108076973A (en) * | 2018-01-15 | 2018-05-29 | 石家庄学院 | A kind of production method of mushroom concentrated strain |
| CN110583371A (en) * | 2019-09-23 | 2019-12-20 | 贵州同辉食用菌发展有限公司 | Fermentation method of mushroom liquid strain with high activity and high stability |
| CN112703967A (en) * | 2020-12-07 | 2021-04-27 | 贵州大学 | Method for preparing wood rot fungus liquid strain by using yellow serofluid and bean dregs |
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Application publication date: 20160309 |